An in-frame deletion mutation in the degron tail of auxin coreceptor IAA2 confers resistance to the herbicide 2,4-D in Sisymbrium orientale.

The natural auxin indole-3-acetic acid (IAA) is a key regulator of many aspects of plant growth and development. Synthetic auxin herbicides such as 2,4-D mimic the effects of IAA by inducing strong auxinic-signaling responses in plants. To determine the mechanism of 2,4-D resistance in a Sisymbrium orientale (Indian hedge mustard) weed population, we performed a transcriptome analysis of 2,4-D-resistant (R) and -susceptible (S) genotypes that revealed an in-frame 27-nucleotide deletion removing nine amino acids in the degron tail (DT) of the auxin coreceptor Aux/IAA2 (SoIAA2). The deletion allele cosegregated with 2,4-D resistance in recombinant inbred lines. Further, this deletion was also detected in several 2,4-D-resistant field populations of this species. Arabidopsis transgenic lines expressing the SoIAA2 mutant allele were resistant to 2,4-D and dicamba. The IAA2-DT deletion reduced binding to TIR1 in vitro with both natural and synthetic auxins, causing reduced association and increased dissociation rates. This mechanism of synthetic auxin herbicide resistance assigns an in planta function to the DT region of this Aux/IAA coreceptor for its role in synthetic auxin binding kinetics and reveals a potential biotechnological approach to produce synthetic auxin-resistant crops using gene-editing.

Current status of community resources and priorities for weed genomics research.

Weeds are attractive models for basic and applied research due to their impacts on agricultural systems and capacity to swiftly adapt in response to anthropogenic selection pressures. Currently, a lack of genomic information precludes research to elucidate the genetic basis of rapid adaptation for important traits like herbicide resistance and stress tolerance and the effect of evolutionary mechanisms on wild populations. The International Weed Genomics Consortium is a collaborative group of scientists focused on developing genomic resources to impact research into sustainable, effective weed control methods and to provide insights about stress tolerance and adaptation to assist crop breeding.

Population structure of ALS-inhibiting herbicide-resistant Sonchus oleraceus in South Australia.

BACKGROUND: Annual sowthistle is a weed that is difficult to control in lentil crops in southern Australia due to a lack of herbicide options, widespread herbicide resistance and prolific production of highly mobile seed. This study investigates herbicide resistance in annual sowthistle in the Mid-North (MN) and Yorke Peninsula (YP) regions of South Australia, identifies and characterizes the mechanisms of acetolactate-synthase (ALS)-inhibitor resistance in this amphidiploid species, and combines this with analyses of population structure and gene flow. RESULTS: ALS-inhibitor-resistant annual sowthistle is widespread across the YP and MN of South Australia and is associated with a variety of Proline-197 mutations of the ALS gene, including leucine, alanine, arginine, serine, threonine and histidine. These mutations were found in different combinations on either of the two copies of the ALS gene. An additional 200 tissue samples were collected from across a single field on the YP and the ALS gene was sequenced for all these individuals. Different ALS-inhibitor resistance profiles were evident between mutation combinations and within mutation combinations, possibly mediated by differing subgenome assortment of the mutations, or altered gene experession of the two ALS homeologs. Population genetics analysis showed evidence of long-distance dispersal, resulting in highly mobile resistance genes, and multiple instances of resistance mutation evolution. CONCLUSIONS: Continuing selection of Sonchus oleraceus populations with ALS-inhibiting herbicides has resulted in the accumulation of additional mutations within the ALS gene. New practices to control herbicide-resistant S. oleraceus should be examined, and control should focus on reducing seed set and dispersal to prevent the spread of emerging cases of resistance. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Point mutations including a novel Pro-197-Phe mutation confer cross-resistance to acetolactate synthase (ALS) inhibiting herbicides in Lactuca serriola in Australia.

BACKGROUND: Control of prickly lettuce has become increasingly difficult for lentil growers in southern Australia because of widespread resistance to common herbicides, a lack of alternative herbicide options and the prolific production of highly mobile seed. This study aimed to quantify acetolactate synthase (ALS)-inhibiting herbicide resistance in the Mid North (MN) and Yorke Peninsula (YP) of South Australia, characterize the resistance mutations present and investigate population structure and gene flow in this species. RESULTS: Resistance was identified in all populations tested, with average survival of 92% to chlorsulfuron and 95% to imazamox + imazapyr. Five different amino acid substitutions were identified at proline 197 of the ALS gene. There was no significant difference in the median lethal dose (LD50 ) between plants with these five different substitutions when treated with metsulfuron-methyl; however, the imidazolinone resistance level was higher in plants with a phenylalanine substitution and lower in plants with a serine. Population structure based on 701 single nucleotide polymorphisms and 271 individuals provided evidence for both independent evolution of the same mutation in different populations, as well as frequent short- to medium-distance dispersal accompanied by occasional long-distance dispersal events. The overall inbreeding coefficient (FIS ) was calculated at 0.5174, indicating an intermediate level of outcrossing despite the cross-pollination experiment showing only low outcrossing. In the structure analyses, most individuals from YP were assigned to a single cluster, whereas most individuals from MN were assigned 50% to each of two clusters, indicating some genetic differences between these two regions, but also evidence for dispersal between them. CONCLUSIONS: Use of imidazolinone herbicides has selected for mutations conferring higher levels of resistance, such as the Pro-197-Phe mutation, and resulted in further spread of resistance in this species. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Non-Mendelian inheritance of gene amplification-based resistance to glyphosate in Hordeum glaucum (barley grass) from South Australia.

BACKGROUND: Hordeum glaucum Steud. is an important grass weed species in South Australia that has evolved resistance to glyphosate. This study investigated the mode of inheritance of glyphosate resistance in this species. RESULTS: Hand-pollination of glyphosate susceptible and resistant populations generated two F1 individuals, selfed to yield F2 progenies. In dose-response experiments, the F2 progenies showed intermediate response between the two parent populations. High variation in EPSPS gene copies was observed among F2 individuals, with some individuals possessing more gene copies than the resistant parent population. No evidence of a Mendelian single-gene pattern of inheritance was observed. CONCLUSION: Inheritance of gene amplification in H. glaucum is non-Mendelian. © 2021 Society of Chemical Industry.

Stability of EPSPS gene copy number in Hordeum glaucum Steud (barley grass) in the presence and absence of glyphosate selection.

BACKGROUND: Gene amplification has been shown to provide resistance to glyphosate in several weed species, including Hordeum glaucum populations in South Australia. The stability of 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene copies in resistant populations in the presence or absence of glyphosate selection has not been determined. RESULTS: Applying glyphosate to a cloned plant resulted in an increase in resistance and EPSPS copy number in the progeny of that plant compared to the untreated clone. The LD50 (herbicide concentration required for 50% mortality) increased by 75% to 79% in the progeny of the treated clones compared to the untreated in both populations (YP-17 and YP-16). EPSPS copy number estimates were higher in treated individuals compared to untreated individuals with an average of seven copies compared to six in YP-16 and 11 compared to six in YP-17. There was a positive correlation (R2  = 0.78) between EPSPS copy number and LD50 of all populations. CONCLUSION: EPSPS gene copy number and resistance to glyphosate increased in H. glaucum populations under glyphosate selection, suggesting the number of EPSPS gene copies present is dependent on glyphosate selection. © 2021 Society of Chemical Industry.

EPSPS gene amplification confers resistance to glyphosate resistant populations of Hordeum glaucum Stued (northern barley grass) in South Australia.

BACKGROUND: Glyphosate failed to control two populations of Hordeum glaucum (northern barley grass) along a fence line and around stockyards near Arthurton on the Yorke Peninsula, South Australia after more than a decade of regular use of glyphosate. These were investigated to confirm resistance and to determine resistance mechanisms. RESULTS: Dose-response experiments confirmed resistance of these populations to glyphosate with resistance levels of 2.8-6.6-fold compared with two susceptible populations. Shikimate assays confirmed resistance to glyphosate with less shikimate accumulation in the resistant populations compared with the susceptible populations. Quantitative PCR of genomic DNA showed increased gene copy number in the resistant populations with 9-11-fold more copies of EPSPS compared with the susceptible populations, suggesting resistance is likely conferred by gene amplification. CONCLUSION: This study identified the first examples of glyphosate resistance in the grass species H. glaucum with resistance associated with EPSPS gene amplification. © 2019 Society of Chemical Industry.

Cross-resistance to diflufenican and picolinafen and its inheritance in oriental mustard (Sisymbrium orientale L.).

BACKGROUND: An oriental mustard population (P40) was identified as resistant to diflufenican by screening at the field rate. As diflufenican and picolinafen both target phytoene desaturase (PDS), cross-resistance to picolinafen was suspected. The mechanism of resistance and its inheritance to diflufenican and picolinafen were investigated. RESULTS: At the lethal dose (LD50 ) level, population P40 was 237-fold more resistant to diflufenican and seven-fold more resistant to picolinafen compared to two susceptible populations. Population P40 also had a significantly higher resistance to diflufenican (237-fold) than a previously described P3 population (143-fold). In addition to the Leu-498-Val mutation in PDS identified in all individuals of the P3 and P40 populations, a Glu-425-Asp mutation was also found in P40. Neither mutation was detected in any individuals of the susceptible population. As the segregation of phenotype and genotype of the F2 individuals fitted the model for a single dominant allele, resistance to both diflufenican and picolinafen is likely encoded on the nuclear genome and is dominant. CONCLUSION: Resistance to diflufenican and picolinafen in the P40 population is likely conferred by Leu-498-Val and Glu-425-Asp mutations in the PDS gene. Inheritance of resistance to these herbicides is managed by a single dominant gene. © 2018 Society of Chemical Industry.

EPSPS gene amplification conferring resistance to glyphosate in windmill grass (Chloris truncata) in Australia.

BACKGROUND: Five glyphosate-resistant populations of Chloris truncata originally collected from New South Wales were compared with one susceptible (S) population from South Australia to confirm glyphosate resistance and elucidate possible mechanisms of resistance. RESULTS: Based on the amounts of glyphosate required to kill 50% of treated plants (LD50 ), glyphosate resistance (GR) was confirmed in five populations of C. truncata (A536, A528, T27, A534 and A535.1). GR plants were 2.4-8.7-fold more resistant and accumulated less shikimate after glyphosate treatment than S plants. There was no difference in glyphosate absorption and translocation between GR and S plants. The EPSPS gene did not contain any point mutation that had previously been associated with resistance to glyphosate. The resistant plants (A528 and A536) contained up to 32-48 more copies of the EPSPS gene than the susceptible plants. CONCLUSION: This study has identified EPSPS gene amplification contributing to glyphosate resistance in C. truncata. In addition, a Glu-91-Ala mutation within EPSPS was identified that may contribute to glyphosate resistance in this species. © 2017 Society of Chemical Industry.

Reduced translocation in 2,4-D-resistant oriental mustard populations (Sisymbrium orientale L.) from Australia.

BACKGROUND: Two oriental mustard populations (P2 and P13) collected from Port Broughton, South Australia were identified as resistant to 2,4-D. The level of resistance, mechanism and the mode of inheritance for 2,4-D resistance in these populations were investigated. RESULTS: Populations P2 and P13 were confirmed to be resistant to 2,4-D at the field rate (600 g a.e. ha-1 ). P2 and P13 were 81- and 67-fold more resistant than the susceptible populations (S1 and S2) at the dose required for 50% mortality (LD50 ), respectively. No predicted amino acid modification was detected in sequences of potential target-site genes (ABP, TIR1 and AFB5). Resistant populations had reduced 2,4-D translocation compared with the susceptible populations, with 77% of [14 C]2,4-D retained in the treated leaf versus 32% at 72 h after treatment. Resistance to 2,4-D is encoded on the nuclear genome and is dominant, as the response to 2,4-D of all F2 individuals were similar to the resistant biotypes. The segregation of F2 phenotypes fitted a 3: 1 (R: S) inheritance model. CONCLUSION: Resistance to 2,4-D in oriental mustard is likely due to reduced translocation of 2,4-D out of the treated leaf. Inheritance of 2,4-D resistance is conferred by a single gene with a high level of dominance. © 2017 Society of Chemical Industry.

The power and potential of genomics in weed biology and management.

There have been previous calls for, and efforts focused on, realizing the power and potential of weed genomics for better understanding of weeds. Sustained advances in genome sequencing and assembly technologies now make it possible for individual research groups to generate reference genomes for multiple weed species at reasonable costs. Here, we present the outcomes from several meetings, discussions, and workshops focused on establishing an International Weed Genomics Consortium (IWGC) for a coordinated international effort in weed genomics. We review the 'state of the art' in genomics and weed genomics, including technologies, applications, and on-going weed genome projects. We also report the outcomes from a workshop and a global survey of the weed science community to identify priority species, key biological questions, and weed management applications that can be addressed through greater availability of, and access to, genomic resources. Major focus areas include the evolution of herbicide resistance and weedy traits, the development of molecular diagnostics, and the identification of novel targets and approaches for weed management. There is increasing interest in, and need for, weed genomics, and the establishment of the IWGC will provide the necessary global platform for communication and coordination of weed genomics research. © 2018 Society of Chemical Industry.

EPSPS gene amplification in glyphosate-resistant Bromus diandrus.

BACKGROUND: Glyphosate is the most widely used herbicide in the world and has been intensively used to control B. diandrus, a problematic weed of crops and pastures in southern Australia. RESULTS: Resistance to glyphosate was identified in two populations of B. diandrus that were nearly fivefold more resistant to glyphosate than wild-type plants. Both populations contained EPSPS gene amplification, with resistant plants having an average of around 20-fold the number of copies of EPSPS compared with susceptible plants. EPSPS expression was also increased in resistant plants of both populations; however, expression levels were not correlated with the number of EPSPS copies. Amplification of only one of the four EPSPS genes present in B. diandus was detected. Investigation into the inheritance of glyphosate resistance found no segregation in the F2 generation. Every individual in the F2 populations contained between three and 30 copies of EPSPS; however, on average they contained fewer copies compared with the parent resistant population. CONCLUSIONS: Glyphosate resistance in B. diandrus is due to EPSPS gene amplification. Resistance is heritable but complex.

Temperature influences the level of glyphosate resistance in barnyardgrass (Echinochloa colona).

BACKGROUND: Echinochloa colona is an important summer-growing weed species in cropping regions of northern Australia that has evolved resistance to glyphosate owing to intensive use of this herbicide in summer fallow. RESULTS: Pot trials conducted at 20 and 30 °C on six E. colona populations showed a significant increase in the level of glyphosate resistance in resistant populations at 30 °C compared with 20 °C. However, there was no influence of growth temperature on glyphosate susceptibility of the sensitive population. Sequencing of the target-site gene (EPSPS) of the six populations identified a mutation at position 106 leading to a change from proline to serine in the most resistant population A533.1 only. EPSPS gene amplification was not detected in any of the resistant populations examined. Examining (14) C-glyphosate uptake on two resistant and one susceptible population showed a twofold increase at 20 °C; however, few differences in glyphosate translocation occurred from the treated leaf to other plant parts between populations or temperatures. CONCLUSION: There is reduced efficacy of glyphosate at high temperatures on resistant E. colona populations, making these populations harder to control in summer.

Inheritance of resistance to 2,4-D and chlorsulfuron in a multiple-resistant population of Sisymbrium orientale.

BACKGROUND: A population of Sisymbrium orientale from South Australia has multiple resistance to auxinic herbicides and inhibitors of acetohydroxyacid synthase (AHAS). Inheritance of resistance to 2,4-D and chlorsulfuron was studied in this population. RESULTS: Crosses were made between seven resistant individuals as pollen donors to seven susceptible individuals. Sixteen F1 individuals from three crosses were identified by their lack of strong epinasty when treated with 200 g 2,4-D ha(-1). These individuals were selfed, and segregation analysis of strong epinasty in the resulting progeny fitted a 3:1 ratio for resistant:susceptible individuals when treated with 200 g 2,4-D ha(-1), as predicted by a single major gene. A detailed dose-response analysis of the F2 populations to 2,4-D confirmed single-gene inheritance. Analysis of segregation to 1 g chlorsulfuron ha(-1), a concentration that kills all susceptible individuals, was unable to determine the mode of inheritance. A detailed dose-response analysis indicated that two genes contributed to chlorsulfuron resistance: a dominant target-site mutation of Pro 197 to Ser and a second gene with dose-dependent dominance. CONCLUSIONS: This population has a single dominant allele conferring 2,4-D resistance, whereas two genes contribute to chlorsulfuron resistance. Single dominant gene inheritance demonstrates that 2,4-D resistance can be readily selected.